이 작품 Odontological 연구 (IFRO) 대학 파리 디드로, 프랑스 국립 건강 연구소 및 의료 연구 (INSERM), 그리고 프랑스 연구소에 의해 투자 되었다.

현재 연구에 사용 된 모든 동물 관리 및 농업 (A-75-06-12)의 프랑스 내각에서 실험 동물의 사용에 대 한 지침에 따라 유지 되었다.
1. 동물 유독 노출
<ol><li>이 프로토콜을 수행 하기 전에 필요한 기관 승인을 하 고 모든 동물 보호 지침을 준수 해야 합니다.</li>
	<li>Ameloblast 분 비 단계 및 ameloblast 성숙 단계에 조사 하는 molecule(s)의 영향을 테스트 하기 위해 다른 실험 그룹의 헌법을 수 있도록 연구 프로토콜의 디자인을 적용 합니다. 여기, 남성 Wistar 쥐의 4 개 그룹 불 (NaF) 조합에서 또는 하지 BPA (그림 1A)2에 그들의 노출에 따라 구성 되었다. 모든 동물 관찰 하 고 하루 65 (그림 1B)에 해 부 했다.</li>
</ol>2. Hemi-Mandibles 성인 쥐에서의 해 부
<ol><li>CO2 질, 선택적으로 시체의 나머지 부분에서 별도 머리를 잘린 다음으로 쥐를 안락사. CO2 전용된 상자16에서 5 분에 쥐를 노출 합니다.</li>
	<li>#11 메스를 사용 하 여 더 낮은 앞 니에 쉽게 접근할 낮은 입술에서 모든 피부를 제거 합니다.</li>
	<li>같은 메스와 약간의 압력 두 더 낮은 앞 니 사이 절 개 하 고는 mandible 두 개의 반쪽으로 분할 됩니다.</li>
	<li>턱, 분리를 메스로 일시적인 악의 공동 잘라내어 hemi mandible 정밀한 핀셋으로 잡고.</li>
	<li>메스와 주변 부드러운 조직 고 모든 근육, 힘 줄, 인 대는 뼈가 완전히 깨끗 한 때까지 스 크레이 퍼를 사용 하 여 제거.</li>
</ol>3. 니17,18 의 격리
<ol><li>신중 하 게 블레이드는 니의 주요 종 축에 평행 하 게 배치 하 여 뼈 떨어져 면도. 자 궁 경부 루프 근처 니 끝까지 니 (근 끝)의 끝 근처 뼈 능선에서 시작 합니다. 절 개 모션 원심 방향에 인접에서 진행 됩니다.</li>
	<li>에 게 첫 번째 컷 distally hemi mandible의 gonion를 제거 하 고 쉽게 자 궁 경부 루프 또는 ameloblasts ( 그림 2에서 레드 라인)의 분 비 단계 손상 없이 니 이륙 허가를 메스로 자 궁 경관 루프.</li>
	<li>어 금 니, 두 번째 아래 두 번째 컷을 확인 하 고 뼈와 고 니의 중간 표면 사이 메스를 삽입. 모든 기저 뼈를 들어올리면는 니 바깥쪽으로 회전 하 고에 나 멜을 피하기 위해 그것을 신중 하 게 벗어 기관 조직 손상 정밀한 핀셋을 사용 하 여.</li>
</ol>4. 마이크로-의 해 부 눈 렌즈 아래에 나 멜 기관
<ol><li>피 펫을 사용 하 여 니에 염 인산 염 버퍼 (PBS 1x)의 200 µ L을 드롭. 고급 핀셋으로 니 위로 향하도록 입술 표면 가져가 라. 메스는 무색 조직의 주황색 부분 사이 표시를 확인 합니다. 이 마크는 나중에 볼 (그림 2)는 기본 백색 반점에 해당 합니다.</li>
	<li>굴 삭 기 또는 해당 도구와 분 비 단계 (무색) ameloblasts, 그리고 성숙 단계 (오렌지) ameloblasts 대 니의 끝을 메스 마크에서에 해당 하는 꼭대기 끝을 메스 마크에서 세포 표면을 다쳤어요.</li>
	<li>이전 설명된19로 전환 단계로 해당 2 mm 조직을 제거 합니다. 사기 질 기관 절 개는 mesenchyme 하 여 오염을 방지 하는 동안에 고 니를 열지 마십시오.</li>
	<li>앞에서 설명한20로 니 (그림 2)의 꼭대기 부분에 바로 위치한 자 궁 경부 루프를 잘라.</li>
	<li>추가 조사 ( 재료의 표참조)에 대 한 10% 포 르 말린 또는 세포 솔루션에 그것을 수집 합니다.</li>
</ol>5. 추가 조사에 대 한 분리 된 사기 질 기관 직물의 수집
<ol><li>200 µ L PBS 1 X 버퍼는 니에 드롭.</li>
	<li>#11 메스 블레이드, 신중 하 게 분리, 버퍼에 점차적으로 치과 상피 세포 별도로 이전 메스 마크의 도움으로 각 단계에 대 한.</li>
	<li>셀 RNA 및 단백질 추출, 단백질 및 RNAs ( 재료의 표참조)의 적 출을 허용 하는 세포 해결책을 사용 하 여 조직을 넣어. 참고: 조직-굴 삭 기는 튜브에 하 고 다음 셀 균질 혼합물을 얻기까지 연 삭 하 여 고품질 RNAs를 준비 합니다. 혼합물은 앞에서 설명한2,,817제조업체의 절차에 따라 RNA 및 단백질 추출에 대 한 준비. 일반적으로, 약 5-10 µ g 총 RNAs 및 150 µ g 총 단백질 각 준비를 획득 했다.</li>
	<li>조직학 분석에 대 한 immunohistochemistry (IHC), 그리고 현장에서 교 잡 (ISH)는 2 h, 그 후에 PBS 1 X에서에서 4 ° C에서 저장소에 대 한 10% 포 르 말린에 셀 레이어를 넣어. 조직 수 포함할 수 왁 스/파라핀 또는 조직에 10 월 냉동된 섹션에 대 한.</li>
	<li>EMP 기사는 니 고급 핀셋으로 걸릴. 공기 (약간의 탈수)에 짧은 노출 후 하얀 점이 사기 질 강화 작용의 개시를 나타내는 니 입술의 가운데 부분 주위에 표시 됩니다. 이 백색 반점 / 해당 하는 전환-초기 성숙 단계의 ameloblasts, 그래서 그것을 사용 표시기로 EMPs21의 추출.</li>
</ol>

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L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Identification+of+CCAAT/enhancer-binding+protein+alpha+as+a+transactivator+of+the+mouse+amelogenin+gene.">Identification of CCAAT/enhancer-binding protein alpha as a transactivator of the mouse amelogenin gene.</a> J Biol Chem. 275, 12273-12280 (2000).</li><li>Nakata, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Establishment+and+characterization+of+a+spontaneously+immortalized+mouse+ameloblast-lineage+cell+line.">Establishment and characterization of a spontaneously immortalized mouse ameloblast-lineage cell line.</a> Biochem Biophys Res Commun. 308, 834-839 (2003).</li><li>Harada, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Establishment+of+ameloblastoma+cell+line,+AM-1.">Establishment of ameloblastoma cell line, AM-1.</a> Journal of oral pathology &amp; medicine: official publication of the International Association of Oral Pathologists and the American Academy of Oral Pathology. 27, 207-212 (1998).</li><li>Jussila, M., Thesleff, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Signaling+networks+regulating+tooth+organogenesis+and+regeneration,+and+the+specification+of+dental+mesenchymal+and+epithelial+cell+lineages.">Signaling networks regulating tooth organogenesis and regeneration, and the specification of dental mesenchymal and epithelial cell lineages.</a> Cold Spring Harb Perspect Biol. 4, a008425 (2012).</li><li>Tucker, A., Sharpe, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+cutting-edge+of+mammalian+development;+how+the+embryo+makes+teeth.">The cutting-edge of mammalian development; how the embryo makes teeth.</a> Nat Rev Genet. 5, 499-508 (2004).</li><li>Vos, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Years+lived+with+disability+(YLDs)+for+1160+sequelae+of+289+diseases+and+injuries+1990-2010:+a+systematic+analysis+for+the+Global+Burden+of+Disease+Study+2010.">Years lived with disability (YLDs) for 1160 sequelae of 289 diseases and injuries 1990-2010: a systematic analysis for the Global Burden of Disease Study 2010.</a> Lancet. 380, 2163-2196 (2012).</li><li>Marcenes, W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Global+burden+of+oral+conditions+in+1990-2010:+a+systematic+analysis.">Global burden of oral conditions in 1990-2010: a systematic analysis.</a> J Dent Res. 92, 592-597 (2013).</li><li>. Dental Caries (Tooth Decay) in Adults (Age 20 to 64) Available from: <a target="_blank" href="https://www.nidcr.nih.gov/DataStatistics/FindDataByTopic/DentalCaries/DentalCariesAdults20to64.htm">https://www.nidcr.nih.gov/DataStatistics/FindDataByTopic/DentalCaries/DentalCariesAdults20to64.htm</a> (2017)</li></ol>

저자는 공개 관심의 없습니다 충돌 있다.

변경 된 ameloblast 활동 또는 방해 ameloblast 확산, 차별화, 그리고 성숙 프로세스 리드 돌이킬 수 없는 사기 질 결함을, 차례로,에 나 멜 결함의 특성은 변경의 이해 도움이 될 수 있습니다. amelogenesis 중 ameloblast 활동입니다. 따라서, 고립 된 사기 질 기관에 학문은 사기 질 결점 든 그들의 근원, 환경 또는 유전을 선도 병 적인 이벤트를 명료 하 게 하는 결정.
이 기술은 원래 Hiller <em...

많은 발달에 나 멜 결함 환경 유독 또는 부적 절 한 생활 스타일1,2,,34에서 발생할 수 있습니다. 이벤트 및 현재 설명한 절차를 사용 하 여 amelogenesis의 분자의 여러 유독 노출의 조기 표식으로 결과 나 멜 결함의 사용을 촉진 것입니다 그리고 건강의 역사를 다시 구성 하는 데 도움이 될 수 있습니다. 때에 나 멜은 synthetized1,2주 기간 동안 각 환자. 에 나 멜 합성 ameloblast 활동5에 따라 네 가지 주요 단계로 나눌 수 있습니다. 첫 번째 단계 regroups 전조 세포 및 pre-ameloblast 확산. 두 번째 단계 동안 차별화 된 ameloblasts에 나 멜 매트릭스 단백질 (EMPs), 주로 amelogenin, enamelin 및 ameloblastin, 마지막에 나 멜의 두께 결정 하는 분 비. 따라서, EMP 합성의 중단에 나 멜의 정량적 결함 이끌어 낸다. 전체에 나 멜 두께의 증 착 후 성숙 단계는 시작 됩니다. 이 단계 동안, 인회석 crystallite 성장 폭과 두께에 최대 96% 무게에 의해 생물 학적 조직에 높은 강화 비율을 도달 하는 나 멜을 수 있습니다. 질적 성숙 단계 이어질 동안 발생 하는 방해 이벤트 결함 사기 질. 마지막으로, ameloblasts 설치류, 염색 라고도 후 성숙의 단계를 입력 하 고 돌이킬 수 없는, 돌이킬 수 없는 (있을 경우)에 나 멜 결함을 만드는 치아 분화 동안 apoptosis를 받 다, 따라서 결함의 잠재적인 회고전 기록 제공 ameloblast 고 강조입니다. 설치류, amelogenesis 그들 amelogenesis의 일반적인 과정을 공부 하기 적합 한 모델을 만드는 그들의 앞 니는 지속적으로 성장 하 고, 까다로움과 이벤트의 유사한 순서를 따른다. 따라서, amelogenesis의 중단에 나 멜 품질 및 수량, 방해 이벤트의 기간에 따라 변경 발생합니다. 그런 의미에서 다이옥신, 납, 그리고 내 분 비 방해 화학 물질 (EDCs) 비스 페 놀 A (BPA), genistein, vinclozolin, 등에 노출에 나 멜 hypomineralizations,12,3 생성 하기 위해 표시 되었습니다. ,6,,78. 비대칭 흰색 불투명 반점이 쥐 태아 기간 출생1후 첫 달 동안 낮은 복용량 BPA 복용량에 노출의 앞 니에 확인 되었다. 이 나 멜 결함 쥐, 그리고 인간 어 금 니 니 hypomineralization (MIH)의 그에 유사한 임상, 구조, 및 생 화 확 적인 특성을 공유합니다. MIH는 최근 설명된 치과 나 멜 병 병 인 아직도 남아 있는 많은 인과 요인에도 불구 하 고 불분명9,10 에9,10,11 가상 데 ,12.
환경 요인으로 인해 또 다른 중요 한에 나 멜 hypomineralization 병리학은 치과 fluorosis (DF), 과도 한 불 소 흡수의 결과 이다 (> 0.1 m g/k g/일)13,14. 불 소의 주요 소스는 식 수를 보충 하거나 자연스럽 게 불 소 강화입니다. 불 소는 또한 종종 치과 카 리에 스를 방지 하기 위해 처방 이지만 예방 복용량만 50% 보다 낮은 독성 하나 (0.05 mg/kg/day). MIH와 DF, 환경 요인에 노출에서 결과로 두 자주 pathologies EDCs2 등 다른 유독 함께 하는 불 소의 hypomineralizing 효과의 potentiation 인해 특성화 하는 데 필요한 공통 기능을 제공할 수 있습니다. 또는 amoxicillin15.
쥐 사기 질 기관의 ameloblasts 다른 분화 단계에서 포함 된 마이크로 해 부는 ameloblast 활동을 방해 하 고에 나 멜 치아 분화 후 진단 결함을 일으킬 수 있는 분자의 행동의 메커니즘을 이해 하는 데 도움이 됩니다. 즉,에 나 멜 진 식과에 나 멜 매트릭스 구성 환경 유독 때문의 변화 특성에 유독, 노출의 역사의 재구성 있으며 공개에 대 한 환경 안전 모니터링을 용이 하 게 건강입니다.

<table>
	<tbody>
		<tr>
			<td>Bisphenol A</td>
			<td>Sigma Aldrich, Saint Louis MO</td>
			<td>239658</td>
			<td></td>
		</tr>
		<tr>
			<td>formalin 10%</td>
			<td>Sigma-Aldrich, Saint Louis, MO</td>
			<td>HT5012</td>
			<td></td>
		</tr>
		<tr>
			<td>Tri-Reagent</td>
			<td>Euromedex, France</td>
			<td>TR118</td>
			<td></td>
		</tr>
		<tr>
			<td>RLT buffer</td>
			<td>Qiagen, Les Ulis, France</td>
			<td>74126</td>
			<td>RNeasy Protect Mini Kit</td>
		</tr>
		<tr>
			<td>Androgen receptor antibody</td>
			<td>Santa Cruz Biotechnology, Santa Cruz, CA)</td>
			<td>sc-816</td>
			<td>rabbit polyclonal antibody</td>
		</tr>
		<tr>
			<td>PBS 10x</td>
			<td>EUOMEDEX</td>
			<td>ET330.A</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium fluoride (NaF)</td>
			<td>Sigma-Aldrich, Saint Louis, MO</td>
			<td>S-1504</td>
			<td></td>
		</tr>
		<tr>
			<td>paraplast regular</td>
			<td>Leica microsystems, Nanterre cedex,&#160;France</td>
			<td>39601006</td>
			<td>called was/parafin in the text</td>
		</tr>
		<tr>
			<td>tissue OCT</td>
			<td>VWR, Fontenay-sous-Bois, France</td>
			<td>411243</td>
			<td></td>
		</tr>
		<tr>
			<td>Extra Fine Bonn Scissors - Straight/8.5 cm</td>
			<td>PHYMEP , Paris, France</td>
			<td>14084-08</td>
			<td></td>
		</tr>
		<tr>
			<td>Handle for Scalpel Blades - 12.5 cm</td>
			<td>PHYMEP, Paris, France</td>
			<td>10035-12</td>
			<td></td>
		</tr>
		<tr>
			<td>Curved Scalpel Blade</td>
			<td>PHYMEP , Paris, France</td>
			<td>10035-20</td>
			<td></td>
		</tr>
		<tr>
			<td>Dissecting Knife - Fine/Straight Tip</td>
			<td>PHYMEP , Paris, France</td>
			<td>10055-12</td>
			<td></td>
		</tr>
		<tr>
			<td>Circle Knife</td>
			<td>PHYMEP, Paris, France</td>
			<td>10059-15</td>
			<td></td>
		</tr>
		<tr>
			<td>scalpel blades n&#176;11 Swann-Morton</td>
			<td>VWR, Fontenay-sous-Bois, France</td>
			<td>233-0024</td>
			<td></td>
		</tr>
		<tr>
			<td>binocular lens</td>
			<td>Leica biosystems, Nanterre cedex,&#160;France</td>
			<td>MZFLIII</td>
			<td></td>
		</tr>
	</tbody>
</table>

micro-dissection of enamel organ from mandibular incisor of rats exposed to environmental toxicants

환경 상태와 생활의 방법에서 발생 하는 나 멜 결함은 그들의 높은 보급 때문에 공중 보건 우려. 이러한 결함 세포 라는 ameloblasts, 사기 질 기관에에 나 멜 합성에 대 한 책임의 변경 된 활동에서 발생할. Amelogenesis, 동안 ameloblasts 확산, 차별화, 그리고 죽음의 이벤트의 구체적이 고 정확한 시퀀스를 따릅니다. 지속적으로 성장 하는 앞 니 쥐 생리 및 병 적인 조건에 ameloblast 활동 및 분화 단계를 공부 하 적당 한 실험 모델입니다. 여기, 마이크로 해 부 이용과 환경에 노출 된 쥐의 사기 질 기관에 안정적이 고 일관 된 방법을 설명 합니다. 마이크로 해 부 치과 epithelia 포함 분 비 및 성숙 단계 ameloblasts 질적 실험 immunohistochemistry 분석 등 현장에서 교 잡, 뿐만 아니라와 같은 정량 분석에 대 한 사용할 수 있는 실시간 정량 Pcr, RNA-seq, 그리고 서 부 럽.

많은 치과 fluorosis12,13, 과도 한 불 소 흡수 때문 환경 조건에서 발생할 수 있습니다 또는 사기 질 hypomineralization와 유사한 MIH 일부 EDCs1, 에 노출 때문에 결함에 나 멜 7,22. 이러한 발달에 나 멜 결함 쥐 (그림 1)1,<...

Watch this Scientific Journal Video about Micro-dissection of Enamel Organ from Mandibular Incisor of Rats Exposed to Environmental Toxicants at JoVE.com

Micro-dissection of Enamel Organ from Mandibular Incisor of Rats Exposed to Environmental Toxicants

에 나 멜 형성과 가능한 변경 이해 ameloblast 활동의 연구가 필요 합니다. 여기, 마이크로 해 부 분 비 및 성숙 단계 ameloblasts 추가 양적 및 질적 실험 절차에 대 한 사용할 수 있는 포함 하는 나 멜 장기 하는 안정적이 고 일관 된 방법을 설명 합니다.

사기 질 기관 이용과 환경에 노출 된 쥐의 악의 니에서의 마이크로 해 부

Enamel defects resulting from environmental conditions and ways of life are public health concerns because of their high prevalence. These defects result ...

micro-dissection-enamel-organ-from-mandibular-incisor-rats-exposed-to

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JoVE Journal

Medicine

10.0K 조회수.  Cordeliers Research Centre. 에 나 멜 형성과 가능한 변경 이해 ameloblast 활동의 연구가 필요 합니다. 여기, 마이크로 해 부 분 비 및 성숙 단계 ameloblasts 추가 양적 및 질적 실험 절차에 대 한 사용할 수 있는 포함 하는 나 멜 장기 하는 안정적이 고 일관 된 방법을 설명 합니다.

비디오: Micro-dissection of Enamel Organ from Mandibular Incisor of Rats Exposed to Environmental Toxicants

Technique of Subnormothermic Ex Vivo Liver Perfusion for the Storage, Assessment, and Repair of Marginal Liver Grafts

The success of liver transplantation has resulted in a dramatic organ shortage. In most transplant regions 20-30% of patients on the waiting list for liver transplantation die without receiving an organ transplant or are delisted for disease progression. One strategy to increase the donor pool is the utilization of marginal grafts, such as fatty livers, grafts from older donors, or donation after cardiac death (DCD). The current preservation technique of cold static storage is only poorly tolerated by marginal livers resulting in significant organ damage. In addition, cold static organ storage does not allow graft assessment or repair prior to transplantation.
These shortcomings of cold static preservation have triggered an interest in warm perfused organ preservation to reduce cold ischemic injury, assess liver grafts during preservation, and explore the opportunity to repair marginal livers prior to transplantation. The optimal pressure and flow conditions, perfusion temperature, composition of the perfusion solution and the need for an oxygen carrier has been controversial in the past.
In spite of promising results in several animal studies, the complexity and the costs have prevented a broader clinical application so far. Recently, with enhanced technology and a better understanding of liver physiology during ex vivo perfusion the outcome of warm liver perfusion has improved and consistently good results can be achieved.
This paper will provide information about liver retrieval, storage techniques, and isolated liver perfusion in pigs. We will illustrate a) the requirements to ensure sufficient oxygen supply to the organ, b) technical considerations about the perfusion machine and the perfusion solution, and c) biochemical aspects of isolated organs.

Technique of Subnormothermic Ex Vivo Liver Perfusion for the Storage, Assessment, and Repair of Marginal Liver Grafts

Marginal grafts, such as fatty livers, grafts from older donors, or livers retrieved after cardiac death (DCD) tolerate conventional, cold static storage only poorly. We developed a novel model of subnormothermic ex vivo liver perfusion for preservation, assessment, and repair of marginal liver grafts prior to transplantation.

Subnormothermic의 기술<em&gt; 전의 VIVO</em&gt; 간 스토리지, 평가를위한 관류 및 한계 간 이식의 수리

The success of liver transplantation has resulted in a dramatic organ shortage. In most transplant regions 20-30% of patients on the waiting list for ...

연구

의학

Fundus Photography as a Convenient Tool to Study Microvascular Responses to Cardiovascular Disease Risk Factors in Epidemiological Studies

The microcirculation consists of blood vessels with diameters less than 150 &#181;m. It makes up a large part of the circulatory system and plays an important role in maintaining cardiovascular health. The retina is a tissue that lines the interior of the eye and it is the only tissue that allows for a non-invasive analysis of the microvasculature. Nowadays, high-quality fundus images can be acquired using digital cameras. Retinal images can be collected in 5 min or less, even without dilatation of the pupils. This unobtrusive and fast procedure for visualizing the microcirculation is attractive to apply in epidemiological studies and to monitor cardiovascular health from early age up to old age.
Systemic diseases that affect the circulation can result in progressive morphological changes in the retinal vasculature. For example, changes in the vessel calibers of retinal arteries and veins have been associated with hypertension, atherosclerosis, and increased risk of stroke and myocardial infarction. The vessel widths are derived using image analysis software and the width of the six largest arteries and veins are summarized in the Central Retinal Arteriolar Equivalent (CRAE) and the Central Retinal Venular Equivalent (CRVE). The latter features have been shown useful to study the impact of modifiable lifestyle and environmental cardiovascular disease risk factors.
The procedures to acquire fundus images and the analysis steps to obtain CRAE and CRVE are described. Coefficients of variation of repeated measures of CRAE and CRVE are less than 2% and within-rater reliability is very high. Using a panel study, the rapid response of the retinal vessel calibers to short-term changes in particulate air pollution, a known risk factor for cardiovascular mortality and morbidity, is reported. In conclusion, retinal imaging is proposed as a convenient and instrumental tool for epidemiological studies to study microvascular responses to cardiovascular disease risk factors.

Retinal image analysis is an unobtrusive procedure for visualizing the microcirculation. The impact of cardiovascular disease risk factors can result in changes of retinal vessel calibers. The procedures to acquire fundus images and the steps for calculating the vessel calibers are described.

편리한 도구로 안저 사진은 역학 연구에서 심혈관 질환의 위험 요인에 대한 미세 혈관 응답을 연구하기 위해

The microcirculation consists of blood vessels with diameters less than 150 &#181;m. It makes up a large part of the circulatory system and plays an ...

Technique of Porcine Liver Procurement and Orthotopic Transplantation using an Active Porto-Caval Shunt

The success of liver transplantation has resulted in a dramatic organ shortage. Each year, a considerable number of patients on the liver transplantation waiting list die without receiving an organ transplant or are delisted due to disease progression. Even after a successful transplantation, rejection and side effects of immunosuppression remain major concerns for graft survival and patient morbidity. 
Experimental animal research has been essential to the success of liver transplantation and still plays a pivotal role in the development of clinical transplantation practice. In particular, the porcine orthotopic liver transplantation model (OLTx) is optimal for clinically oriented research for its close resemblance to human size, anatomy, and physiology. 
Decompression of intestinal congestion during the anhepatic phase of porcine OLTx is important to guarantee reliable animal survival. The use of an active porto-caval-jugular shunt achieves excellent intestinal decompression. The system can be used for short-term as well as long-term survival experiments. The following protocol contains all technical information for a stable and reproducible liver transplantation model in pigs including post-operative animal care.

Experimental animal research plays a pivotal role in the development of clinical transplantation practice. The porcine orthotopic liver transplantation model (OLTx) closely resembles human conditions and is frequently used in clinically oriented research. The following protocol contains all information for a reliable porcine OLTx model using an active porto-caval-jugular shunt.

활성 포르토 대정맥 션트를 사용하여 돼지 간 조달 및 동소 이식 이식의 기술

The success of liver transplantation has resulted in a dramatic organ shortage. Each year, a considerable number of patients on the liver transplantation ...

Automated Measurement of Pulmonary Emphysema and Small Airway Remodeling in Cigarette Smoke-exposed Mice

COPD is projected to be the third most common cause of mortality world-wide by 2020(1). Animal models of COPD are used to identify molecules that contribute to the disease process and to test the efficacy of novel therapies for COPD. Researchers use a number of models of COPD employing different species including rodents, guinea-pigs, rabbits, and dogs(2). However, the most widely-used model is that in which mice are exposed to cigarette smoke. Mice are an especially useful species in which to model COPD because their genome can readily be manipulated to generate animals that are either deficient in, or over-express individual proteins. Studies of gene-targeted mice that have been exposed to cigarette smoke have provided valuable information about the contributions of individual molecules to different lung pathologies in COPD(3-5). Most studies have focused on pathways involved in emphysema development which contributes to the airflow obstruction that is characteristic of COPD. However, small airway fibrosis also contributes significantly to airflow obstruction in human COPD patients(6), but much less is known about the pathogenesis of this lesion in smoke-exposed animals. To address this knowledge gap, this protocol quantifies both emphysema development and small airway fibrosis in smoke-exposed mice. This protocol exposes mice to CS using a whole-body exposure technique, then measures respiratory mechanics in the mice, inflates the lungs of mice to a standard pressure, and fixes the lungs in formalin. The researcher then stains the lung sections with either Gill&#8217;s stain to measure the mean alveolar chord length (as a readout of emphysema severity) or Masson&#8217;s trichrome stain to measure deposition of extracellular matrix (ECM) proteins around small airways (as a readout of small airway fibrosis). Studies of the effects of molecular pathways on both of these lung pathologies will lead to a better understanding of the pathogenesis of COPD.

The goal of this protocol is to provide automated methods to quantify chronic lung pathologies in a murine model of COPD. The protocol includes exposing mice to cigarette smoke (CS), measuring pulmonary function, inflating the lungs, and using morphometry methods to measure emphysema and small airway remodeling in mice.

담배 연기에 노출 된 마우스의 폐 폐기종 및 작은기도 리모델링 자동 측정

COPD is projected to be the third most common cause of mortality world-wide by 2020(1).  Animal models of COPD are used to identify molecules that ...

Cardiac Catheterization in Mice to Measure the Pressure Volume Relationship: Investigating the Bowditch Effect

Animal models that mimic human cardiac disorders have been created to test potential therapeutic strategies. A key component to evaluating these strategies is to examine their effects on heart function. There are several techniques to measure in vivo cardiac mechanics (e.g., echocardiography, pressure/volume relations, etc.). Compared to echocardiography, real-time left ventricular (LV) pressure/volume analysis via catheterization is more precise and insightful in assessing LV function. Additionally, LV pressure/volume analysis provides the ability to instantaneously record changes during manipulations of contractility (e.g., &#946;-adrenergic stimulation) and pathological insults (e.g., ischemia/reperfusion injury). In addition to the maximum (+dP/dt) and minimum (-dP/dt) rate of pressure change in the LV, an accurate assessment of LV function via several load-independent indexes (e.g., end systolic pressure volume relationship and preload recruitable stroke work) can be attained. Heart rate has a significant effect on LV contractility such that an increase in the heart rate is the primary mechanism to increase cardiac output (i.e., Bowditch effect). Thus, when comparing hemodynamics between experimental groups, it is necessary to have similar heart rates. Furthermore, a hallmark of many cardiomyopathy models is a decrease in contractile reserve (i.e., decreased Bowditch effect). Consequently, vital information can be obtained by determining the effects of increasing heart rate on contractility. Our and others data has demonstrated that the neuronal nitric oxide synthase (NOS1) knockout mouse has decreased contractility. Here we describe the procedure of measuring LV pressure/volume with increasing heart rates using the NOS1 knockout mouse model.

This article describes the measurement of murine left ventricular function via pressure/volume analysis at different heart rates.

Bowditch 효과 조사 : 압력 볼륨 관계를 측정하기 위해 마우스의 심장 도관

Animal models that mimic human cardiac disorders have been created to test potential therapeutic strategies. A key component to evaluating these ...

Murine Prostate Micro-dissection and Surgical Castration

Mouse models are used extensively to study prostate cancer and other diseases. The mouse is an excellent model with which to study the prostate and has been used as a surrogate for discoveries in human prostate development and disease. Prostate micro-dissection allows consistent study of lobe-specific prostate anatomy, histology, and cellular characteristics in the absence of contamination of other tissues. Testosterone affects prostate development and disease. Androgen deprivation therapy is a common treatment for prostate cancer patients, but many prostate tumors become castration-resistant. Surgical castration of mouse models allows for the study of castration resistance and other facets of hormonal biology on the prostate. This procedure can be coupled with testosterone reintroduction, or hormonal regeneration of the prostate, a powerful method to study stem cell lineages in the prostate. Together, prostate micro-dissection and surgical castration opens up a multitude of opportunities for robust and consistent research of prostate development and disease. This manuscript describes the protocols for prostate micro-dissection and surgical castration in the laboratory mouse.

This manuscript describes the protocols for prostate micro-dissection and surgical castration in the laboratory mouse. We also depict representative results produced by these protocols. Finally, we discuss the advantages and utilization of these protocols.

쥐의 전립선 마이크로 해부 및 수술 거세

Mouse models are used extensively to study prostate cancer and other diseases. The mouse is an excellent model with which to study the prostate and has ...

Generation of Organ-conditioned Media and Applications for Studying Organ-specific Influences on Breast Cancer Metastatic Behavior

Breast cancer preferentially metastasizes to the lymph node, bone, lung, brain and liver in breast cancer patients. Previous research efforts have focused on identifying factors inherent to breast cancer cells that are responsible for this observed metastatic pattern (termed organ tropism), however much less is known about factors present within specific organs that contribute to this process. This is in part because of a lack of in vitro model systems that accurately recapitulate the organ microenvironment. To address this, an ex vivo model system has been established that allows for the study of soluble factors present within different organ microenvironments. This model consists of generating conditioned media from organs (lymph node, bone, lung, and brain) isolated from normal athymic nude mice. The model system has been validated by demonstrating that different breast cancer cell lines display cell-line specific and organ-specific malignant behavior in response to organ-conditioned media that corresponds to their in vivo metastatic potential. This model system can be used to identify and evaluate specific organ-derived soluble factors that may play a role in the metastatic behavior of breast and other types of cancer cells, including influences on growth, migration, stem-like behavior, and gene expression, as well as the identification of potential new therapeutic targets for cancer. This is the first ex vivo model system that can be used to study organ-specific metastatic behavior in detail and evaluate the role of specific organ-derived soluble factors in driving the process of cancer metastasis.

This manuscript describes an ex vivo model system comprised of organ-conditioned media derived from the lymph node, bone, lung, and brain of mice. This model system can be used to identify and study organ-derived soluble factors and their effects on the organ tropism and metastatic behavior of cancer cells.

유방암 전이성 행동에 장기 별 영향 유학을위한 장기 에어컨 미디어 및 응용 프로그램의 생성

Breast cancer preferentially metastasizes to the lymph node, bone, lung, brain and liver in breast cancer patients. Previous research efforts have focused ...

Isolation and Cultivation of Mandibular Bone Marrow Mesenchymal Stem Cells in Rats

Here we present an efficient method for isolating and culturing mandibular bone marrow mesenchymal stem cells (mBMSCs) in vitro to rapidly obtain numerous high-quality cells for experimental requirements. mBMSCs could be widely used in therapeutic applications as tissue engineering cells in case of craniofacial diseases and cranio-maxillofacial regeneration in the future due to the excellent self-renewal ability and multi-lineage differentiation potential. Therefore, it is important to obtain mBMSCs in large numbers.
In this study, bone marrow was flushed from the mandible and primary mBMSCs were isolated through whole bone marrow adherent cultivation. Furthermore, CD29+CD90+CD45&#8722; mBMSCs were purified through fluorescent cell sorting. The second generation of purified mBMSCs were used for further study and displayed potential in differentiating into osteoblasts, adipocytes, and chondrocytes. Utilizing this in vitro model, one can obtain a high number of proliferative mBMSCs, which may facilitate the study of the biological characteristics, the subsequent reaction to the microenvironment, and other applications of mBMSCs.

This article presents a method that combines whole bone marrow adherence and flow cytometry sorting for isolating, cultivating, sorting, and identifying bone marrow mesenchymal stem cells from rat mandibles.

쥐에 있는 간골 골수 관진 줄기 세포의 격리 그리고 경작

Here we present an efficient method for isolating and culturing mandibular bone marrow mesenchymal stem cells (mBMSCs) in vitro to rapidly obtain numerous ...

Operating Transverse Aortic Constriction with Absorbable Suture to Obtain Transient Myocardial Hypertrophy

Based on twice transverse aortic constrictions (TACs) in mice, it is proved that myocardial hypertrophic preconditioning (MHP) could attenuate cardiomyocyte hypertrophy and slow down progression to heart failure. For novices, however, the MHP model is usually quite difficult to establish because of the technical obstacles in ventilator operation, opening the chest repeatedly, and bleeding caused by debanding. To facilitate this model, to increase the surgical success rate and to reduce the incidence of bleeding, we switched to absorbable sutures for the first TAC combing with a ventilator-free technique. Using a 2-week absorbable suture, we demonstrated that this procedure could cause significant myocardial hypertrophy in 2 weeks; and 4 weeks after surgery, myocardial hypertrophy was almost completely regressed to the baseline. Using this protocol, the operators could master the MHP model easily with a lower operation mortality.

This protocol presents an improved method to obtain transient myocardial hypertrophy with absorbable suture, simulating left ventricular hypertrophy decrease after removing pressure overload. It could be valuable for the studies on myocardial hypertrophic preconditioning.

과도 심근 비대증을 얻기 위해 흡수 가능한 봉합사와 횡단 대동맥 수축 을 작동

Based on twice transverse aortic constrictions (TACs) in mice, it is proved that myocardial hypertrophic preconditioning (MHP) could attenuate ...

3D Imaging of PDL Collagen Fibers during Orthodontic Tooth Movement in Mandibular Murine Model

Orthodontic tooth movement is a complex biological process of altered soft and hard tissue remodeling as a result of external forces. In order to understand these complex remodeling processes, it is critical to study the tooth and periodontal tissues within their 3D context and therefore minimize any sectioning and tissue artefacts. Mouse models are often utilized in developmental and structural biology, as well as in biomechanics due to their small size, high metabolic rate, genetics and ease of handling. In principle this also makes them excellent models for dental related studies. However, a major impediment is their small tooth size, the molars in particular. This paper is aimed at providing a step by step protocol for generating orthodontic tooth movement and two methods for 3D imaging of the periodontal ligament fibrous component of a mouse mandibular molar. The first method presented is based on a micro-CT setup enabling phase enhancement&#160;imaging of fresh collagen tissues. The second method is a bone clearing method using ethyl cinnamate that enables imaging through the bone without sectioning and preserves endogenous fluorescence. Combining this clearing method with reporter mice like Flk1-Cre;TdTomato provided a first of its kind opportunity to image the 3D vasculature in the PDL and alveolar bone.

We present a protocol for generating orthodontic tooth movement in mice and methods for 3D visualization of the collagen fibers and blood vessels of periodontal ligament without sectioning.